Abstract: Targeting the unlimited self-renewal capability of leukemic stem cells (LSCs) represents a new promising strategy for leukemia therapy. However, only a small number of genes/mutations have been identified to confer this feature to LSCs, which has significantly hampered our understanding of the underlying mechanism(s) for LSC self-renewal and our ability to design effective inhibitory strategies. Using a novel retroviral insertional mutagenesis strategy, we have discovered and confirmed that Set binding protein 1 (Setbp1), an AT-hook transcription factor gene, once overexpressed could confer unlimited self-renewal capability to normal mouse bone marrow progenitor cells and cause their immortalization in culture. Transplantation of Setbp1-expressing cells into lethally-irradiated congenic recipients could also lead to the development of myeloid leukemias. These oncogenic effects of Setbp1 are supported by the identification of its downstream targets. The relevance of SETBP1 activation to human leukemia development is also indicated by a previous study showing its overexpression in a significant fraction (about 8%) of human acute myeloid leukemia cells. Based on these findings, we hypothesize that Setbp1 is a novel oncogene capable of conferring unlimited self-renewal capability to LSCs of myeloid leukemias. Three specific aims have been designed to test this hypothesis. Aim 1 will establish the leukemogenic potential of Setbp1 using a mouse bone marrow transduction and transplantation approach. Taking advantage of insertional mutagenesis by the Setbp1-expressing virus in this system, we will also identify mutations that could cooperate with Setbp1 in leukemia induction by cloning retroviral integrations present in the developed leukemias. In Aim 2, we will determine the types of hematopoietic progenitors that are susceptible to Setbp1-induced self-renewal. Functional establishment of critical downstream targets and protein domains of Setbp1 will also be employed to reveal the possible molecular mechanisms underlying Setbp1-induced self-renewal. Aim 3 will explore the cooperation between Setbp1 and BCR/ABL in the generation and maintenance of LSCs in a mouse model of chronic myeloid leukemia (CML) myeloid blast crisis. CML patient samples will also be examined to establish the potential human disease relevance of this cooperation. The expected results of our proposed studies will produce new insights into the mechanism(s) conferring unlimited self-renewal capability to LSCs in myeloid leukemias. Our study will also test whether targeting LSC self-renewal is efficient in treating CML blast crisis for which no effective treatments are currently available.